Search Thermo Fisher Scientific
Search Thermo Fisher Scientific
Catalog Number | Quantity | Product Type |
---|---|---|
12183020 | 10 Preps | RNA Mini Kit |
12183018A | 50 Preps | RNA Mini Kit |
12183025 | 250 Preps | RNA Mini Kit |
A29839 | 50 Cartridges | RNA Mini Columns |
12183026 | 50 Cartridges | Homogenizer |
Extra-large binding capacity enables rapid RNA purification using standard laboratory equipment
The PureLink RNA Mini Kit provides rapid purification of total RNA from a wide range of cells and tissue types to yield up to 1000 μg of purified RNA from a single extraction (see Figure 1). High-quality total RNA can be obtained from mini- to midi-prep amounts of starting material with just trace amounts of residual genomic DNA contamination. The extra-large binding capacity enables one kit to handle most RNA isolation needs in a quick 20 minute protocol without the need for special sample processing instruments.
Easy, optional, on-column DNase treatment for sensitive applications
The PureLink RNA Mini Kit columns are highly efficient for isolating high quality total RNA while removing the majority of genomic DNA. In general, most applications requiring RNA from animal tissue or mammalian cell lines do not require additional DNase treatment. However, some applications such as gene expression analysis by qRT-PCR without intron-spanning primers or working with samples from organisms with very small or no introns may require more complete removal of residual contaminating DNA. The PureLink DNase Set (12185010) allows for convenient on-column digestion of DNA during the RNA isolation protocol. Treating with DNase while 'on-column' is easier and allows higher RNA recovery than treating with DNase after the RNA has been isolated. The PureLink DNase Set can also be used to remove residual DNA from RNA that has been previously purified. Both the 'on-column' and post-RNA purification workflows are options available with the PureLink DNase Set.
Simplified RNA purification
The PureLink RNA Mini Kit utilizes guanidine-isothiocyanate lysis buffer to protect the RNA during the isolation steps. Special RNase-free reagents combined with certified RNase-free silica membranes in a unique column configuration allow for a safe and easy procedure that can typically be completed in less than 20 minutes without the need of hazardous phenol/chloroform extraction, CsCl centrifugation, or LiCl or alcohol precipitation.
The PureLink RNA Mini Kit is recommended for use with the Homogenizer (Cat. No. 12183026), designed to homogenize cell or tissue lysates via centrifugation, prior to nucleic acid purification. The Homogenizer is especially effective for clarifying particulates from plant tissues.
The presence of ethanol or salt in the purified RNA can inhibit downstream enzymatic reactions. Ensure that you are using the correct order of wash buffers in the kit for washing, and that Wash Buffer II is discarded in the flow-through. Place the spin cartridge into the wash tube and centrifuge the spin cartridge at maximum speed for 2-3 minutes to completely dry the cartridge.
The RNA could have been contaminated with RNase. Ensure that you are using RNase-free equipment and change gloves frequently. Improper handling can also result in RNA degradation. Ensure samples are processed immediately, and that the lysis is performed quickly after adding the lysis buffer. Lastly, tissues rich in RNase (such as rat pancreas) may require the addition of RNase inhibitors or inactivators to protect the RNA from degradation, or a larger volume of lysis buffer.
Low RNA yield can occur due to the following:
- Incomplete lysis and homogenization: ensure that 10 µL of 2-mercaptoethanol was added per milliliter of lysis buffer, perform all steps at room temperature, decrease the amount of starting material used, use proper homogenization methods, and/or cut tissue samples into smaller pieces to ensure complete tissue immersion in the lysis buffer.
- Poor quality starting material: use fresh samples and process immediately after collection.
- Ethanol may not have been added to Wash Buffer II.
- Incorrect elution conditions may have been used: Add RNase-free water and incubate for 1 minute before centrifugation, following the recommendations for elution in the manual. You can also perform a second elution step to recover more RNA.
Incomplete homogenization or dispersal of precipitate after ethanol addition can lead to clogging of the RNA spin cartridge. Clear the homogenate and remove any particulate or viscous material by centrifugation. Completely disperse any precipitate that forms after adding ethanol to the homogenate. Load only the supernatant onto the RNA spin cartridge to avoid clogging.
The PureLink RNA Mini Kit provides rapid column-based purification of total RNA, without organic lysis (phenol/chloroform). You can obtain up to 1,000 µg of purified RNA from a single extraction. The RiboPure RNA Purification Kits combine a phenol/guanidine thiocyanate solution with a glass-fiber filter purification method.
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